RET signalling provides tumorigenic mechanism and tissue specificity for AIP-related somatotrophinomas

International audienceIt is unclear how loss-of-function germline mutations in the widely-expressed co-chaperone AIP , result in young-onset growth hormone secreting pituitary tumours. The RET receptor, uniquely co-expressed in somatotrophs with PIT1, induces apoptosis when unliganded, while RET supports cell survival when it is bound to its ligand. We demonstrate that at the plasma membrane, AIP is required to form a complex with monomeric-intracellular-RET, caspase-3 and PKCδ resulting in PIT1/CDKN2A-ARF/p53-apoptosis pathway activation. AIP-deficiency blocks RET/caspase-3/PKCδ activation preventing PIT1 accumulation and apoptosis. The presence or lack of the inhibitory effect on RET-induced apoptosis separated pathogenic AIP variants from non-pathogenic ones. We used virogenomics in neonatal rats to demonstrate the effect of mutant AIP protein on the RET apoptotic pathway in vivo. In adult male rats altered AIP induces elevated IGF-1 and gigantism, with pituitary hyperplasia through blocking the RET-apoptotic pathway. In females, pituitary hyperplasia is induced but IGF-1 rise and gigantism are blunted by puberty. Somatotroph adenomas from pituitary-specific Aip -knockout mice overexpress the RET-ligand GDNF, therefore, upregulating the survival pathway. Somatotroph adenomas from patients with or without AIP mutation abundantly express GDNF, but AIP -mutated tissues have less CDKN2A-ARF expression. Our findings explain the tissue-specific mechanism of AIP-induced somatotrophinomas and provide a previously unknown tumorigenic mechanism, opening treatment avenues for AIP -related tumours

The syndrome of central hypothyroidism and macroorchidism: IGSF1 controls TRHR and FSHB expression by differential modulation of pituitary TGFβ and Activin pathways

IGSF1 (Immunoglobulin Superfamily 1) gene defects cause central hypothyroidism and macroorchidism. However, the pathogenic mechanisms of the disease remain unclear. Based on a patient with a full deletion of IGSF1 clinically followed from neonate to adulthood, we investigated a common pituitary origin for hypothyroidism and macroorchidism, and the role of IGSF1 as regulator of pituitary hormone secretion. The patient showed congenital central hypothyroidism with reduced TSH biopotency, over-secretion of FSH at neonatal minipuberty and macroorchidism from 3 years of age. His markedly elevated inhibin B was unable to inhibit FSH secretion, indicating a status of pituitary inhibin B resistance. We show here that IGSF1 is expressed both in thyrotropes and gonadotropes of the pituitary and in Leydig and germ cells in the testes, but at very low levels in Sertoli cells. Furthermore, IGSF1 stimulates transcription of the thyrotropin-releasing hormone receptor (TRHR) by negative modulation of the TGFβ1-Smad signaling pathway, and enhances the synthesis and biopotency of TSH, the hormone secreted by thyrotropes. By contrast, IGSF1 strongly down-regulates the activin-Smad pathway, leading to reduced expression of FSHB, the hormone secreted by gonadotropes. In conclusion, two relevant molecular mechanisms linked to central hypothyroidism and macroorchidism in IGSF1 deficiency are identified, revealing IGSF1 as an important regulator of TGFβ/Activin pathways in the pituitary

Generation of Immortal Cell Lines from the Adult Pituitary: Role of cAMP on Differentiation of SOX2-Expressing Progenitor Cells to Mature Gonadotropes

The pituitary is a complex endocrine tissue composed of a number of unique cell types distinguished by the expression and secretion of specific hormones, which in turn control critical components of overall physiology. The basic function of these cells is understood; however, the molecular events involved in their hormonal regulation are not yet fully defined. While previously established cell lines have provided much insight into these regulatory mechanisms, the availability of representative cell lines from each cell lineage is limited, and currently none are derived from adult pituitary. We have therefore used retroviral transfer of SV40 T-antigen to mass immortalize primary pituitary cell culture from an adult mouse. We have generated 19 mixed cell cultures that contain cells from pituitary cell lineages, as determined by RT-PCR analysis and immunocytochemistry for specific hormones. Some lines expressed markers associated with multipotent adult progenitor cells or transit-amplifying cells, including SOX2, nestin, S100, and SOX9. The progenitor lines were exposed to an adenylate cyclase activator, forskolin, over 7 days and were induced to differentiate to a more mature gonadotrope cell, expressing significant levels of α-subunit, LHβ, and FSHβ mRNAs. Additionally, clonal populations of differentiated gonadotropes were exposed to 30 nM gonadotropin-releasing hormone and responded appropriately with a significant increase in α-subunit and LHβ transcription. Further, exposure of the lines to a pulse paradigm of GnRH, in combination with 17β-estradiol and dexamethasone, significantly increased GnRH receptor mRNA levels. This array of adult-derived pituitary cell models will be valuable for both studies of progenitor cell characteristics and modulation, and the molecular analysis of individual pituitary cell lineages

A GRFa2/Prop1/Stem (GPS) Cell Niche in the Pituitary

BACKGROUND: The adult endocrine pituitary is known to host several hormone-producing cells regulating major physiological processes during life. Some candidates to progenitor/stem cells have been proposed. However, not much is known about pituitary cell renewal throughout life and its homeostatic regulation during specific physiological changes, such as puberty or pregnancy, or in pathological conditions such as tumor development. PRINCIPAL FINDINGS: We have identified in rodents and humans a niche of non-endocrine cells characterized by the expression of GFRa2, a Ret co-receptor for Neurturin. These cells also express b-Catenin and E-cadherin in an oriented manner suggesting a planar polarity organization for the niche. In addition, cells in the niche uniquely express the pituitary-specific transcription factor Prop1, as well as known progenitor/stem markers such as Sox2, Sox9 and Oct4. Half of these GPS (GFRa2/Prop1/Stem) cells express S-100 whereas surrounding elongated cells in contact with GPS cells express Vimentin. GFRa2+-cells form non-endocrine spheroids in culture. These spheroids can be differentiated to hormone-producing cells or neurons outlining the neuroectoderm potential of these progenitors. In vivo, GPSs cells display slow proliferation after birth, retain BrdU label and show long telomeres in its nuclei, indicating progenitor/stem cell properties in vivo. SIGNIFICANCE: Our results suggest the presence in the adult pituitary of a specific niche of cells characterized by the expression of GFRa2, the pituitary-specific protein Prop1 and stem cell markers. These GPS cells are able to produce different hormone-producing and neuron-like cells and they may therefore contribute to postnatal pituitary homeostasis. Indeed, the relative abundance of GPS numbers is altered in Cdk4-deficient mice, a model of hypopituitarism induced by the lack of this cyclin-dependent kinase. Thus, GPS cells may display functional relevance in the physiological expansion of the pituitary gland throughout life as well as protection from pituitary disease

Identification of novel pathways involved in the pathogenesis of human adamantinomatous craniopharyngioma

Activating mutations in the gene encoding β-catenin have been identified in the paediatric form of human craniopharyngioma (adamantinomatous craniopharyngioma, ACP), a histologically benign but aggressive pituitary tumour accounting for up to 10% of paediatric intracranial tumours. Recently, we generated an ACP mouse model and revealed that, as in human ACP, nucleocytoplasmic accumulation of β-catenin (β-catnc) and over-activation of the Wnt/β-catenin pathway occurs only in a very small proportion of cells, which form clusters. Here, combining mouse genetics, fluorescence labelling and flow-sorting techniques, we have isolated these cells from tumorigenic mouse pituitaries and shown that the β-catnc cells are enriched for colony-forming cells when cultured in stem cell-promoting media, and have longer telomeres, indicating shared properties with normal pituitary progenitors/stem cells (PSCs). Global gene profiling analysis has revealed that these β-catnc cells express high levels of secreted mitogenic signals, such as members of the SHH, BMP and FGF family, in addition to several chemokines and their receptors, suggesting an important autocrine/paracrine role of these cells in the pathogenesis of ACP and a reciprocal communication with their environment. Finally, we highlight the clinical relevance of these findings by showing that these pathways are also up-regulated in the β-catnc cell clusters identified in human ACP. As well as providing further support to the concept that pituitary stem cells may play an important role in the oncogenesis of human ACP, our data reveal novel disease biomarkers and potential pharmacological targets for the treatment of these devastating childhood tumours.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-012-0957-9) contains supplementary material, which is available to authorized users

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Contribución al estudio clínico de la estafilococcia generalizada : Tesis presentada para optar al título de doctor en medicina

A la cabeza de portada: Universidad Nacional de Buenos Aires. Facultad de Ciencias Médicas. - Incluye nómina de Catedráticos y Asignaturas. Tesis con dedicatoria